Stroke is a major health problem worldwide. It is the 4th leading cause of death in Singapore. For those stroke patients who survive, most are likely to be left disabled dud in need of significant rehabilitation. Stroke generates a greater disability impact than any other medical condition and has a huge impact on both the family and society.
There are two types of stroke: ischemic and hemorrhagic. Ischemic stroke, which accounts for more than 80% of all stroke incidences, is attributed by the atherosclerotic occlusion or embolism within an artery, commonly the middle cerebral artery. Focal ischemic stroke with sufficient severity and duration leads to infarction and persistent neurological dysfunction.
Reperfusion is the only potent therapy for acute ischemic stroke. Reperfusion aims to lyse the thrombus with recombinant tissue-type plasminogen activator (rt-PA) or other mechanical devices. Reperfusion therapy is best given within a very narrow time window (<4.5 hours after stroke onset). After that, reperfusion greatly increases cerebral edema and the risk of hemorrhagic transformation which are mainly caused by vascular damage. As very few stroke patients can arrive at hospitals and be diagnosed within this time frame, less than 5% of patients receive reperfusion therapy. Therefore, the focus of acute stroke treatment is to extend the therapeutic time window. However, numerous attempts from both pharmaceutical companies and stroke research community have failed to achieve this goal.
Transient receptor potential melastatin 4 (TRPM4) (see, for example, SEQ ID NOs: 11-13) is a voltage-dependent, non-selective monovalent cation channel. It is impermeable to Ca2+, activated by elevated cytosolic Ca2+, and modulated by ATP [Vennekens R, Nilius B., Handb Exp Pharmacol, 269-85 (2007b)]. TRPM4 belongs to the mammalian TRP superfamily. TRPM4 and TRPM5 are unique because they only conduct monovalent cations, whereas most other TRP channels are permeable to both monovalent and divalent ions. TRPM4 is important for the function of immune cells, including dendritic, mast, and T cells [e.g., Vennekens R, et al., Nat Immunol 8, 312-20 (2007a)]. When activated, TRPM4 can depolarize the membrane potential and regulate Ca2+ homeostasis by decreasing the driving force for Ca2+ entry. Gain-of-function mutations in TRPM4 are associated with familial heart disease [Kruse M, et al., J Clin Invest, 119, 2737-44 (2009)]. TRPM4 also participates in the pathophysiology of spinal cord injury (SCI) and experimental autoimmune encephalomyelitis (EAE) [Gerzanich V, et al., Nat Med, 15, 185-91 (2009); Schattling B, et al., Nat Med, 18, 1805-11 (2012)]. Ectopic expression of TRPM4 has been found in capillaries after SCI and in neurons after EAE. Activation of TRPM4 in SCI and EAE results in unchecked ion influx and subsequently leads to oncotic cell death.
Cerebral edema following brain injury is bound to cell death. Edema resulting from ischemic stroke leads to tissue damage and worsens neurological functions. Recently, upregulation of the non-selective cation channel NCCa-ATP was observed in neurovascular cells, including astrocytes, neurons, and vascular endothelia, after ischemic stroke [Simard J M, et al., Nat Med, 12, 433-40 (2006)]. Enhanced NCCa-ATP current can lead to unchecked Na+ entry, subsequently oncotic cell death, and is believed to cause brain edema [Kahle K T, et al., Physiology (Bethesda), 24, 257-65 (2009)]. The current exhibits many properties similar to those of TRPM4; including a smaller single-channel conductance, permeability to Na+ and Cs+, and activation by intracellular Ca2+ [Simard J M, et al., Nat Med, 12, 433-40 (2006)].
The NCCa-ATP current is also involved in other central nervous system injuries, including traumatic brain injury, spinal cord injury, and subarachnoid hemorrhage [Simard J M, et al., J Neurosurg, 113, 622-9 (2010)]. However, studies of NCCa-ATP channel in stroke have mainly focused on the sulfonylurea receptor-1 (SUR1), an auxiliary subunit of KATP channels [Simard J M, et al., Nat Med, 12, 433-40 (2006)]. After CNS injury, SUR1 has been found upregulated in neurons, astrocytes, and endothelial cells, and the expression is not coupled with KATP functions. There is evidence that SUR1 can associate with TRPM4, and it is believed that blocking SUR1 with a sulfonylurea such as glibenclimide could inhibit the SUR1/TRPM4 channel and salvage brain tissues after injury [Walcott B P, et al., Neurotherapeutics 9:65-72 (2012)]. However, TRPM4 homomers are not sensitive to glibenclamide and there are contradicting reports on whether SUR1 binds TRPM4 directly and whether glibenclimide has a therapeutic effect on stroke [Sala-Rabanal M, et al., J Biol Chem 287:8746-56 (2012); Woo S K, et al., J Biol Chem M112.428219 (2012); Favilla C G, et al., Stroke 42:710-5 (2011); Kunte H, et al., Ann Neurol 72:799-806 (2012)].
After the acute stage, the focus of stroke therapy is to promote angiogenesis and neurogenesis, aiming to improve functional recovery and quality of life of patients. As neurological deficits are severe and always lead to disability among stroke survivors, there is also an urgent need to improve current chronic treatment for stroke recovery.
In view of the above deficiencies; it is desirable to provide a method for extending the window for acute therapy by reducing cerebral edema, and improve current therapy for stroke recovery.
The role of TRPM4 after ischemic stroke is unclear. Rat permanent and transient middle cerebral artery occlusion models (MCAO) were used to investigate the expression and functions of TRPM4 in ischemic stroke and the possibility of inhibiting TRPM4 was tested.